Next-generation Sequencing Advancements in the Diagnostic Lab

An increasing number of clinical laboratories are adopting next-generation sequencing (NGS) in diagnostics, both due to the declining costs of sequencing technologies and the robust biomarker data that can be generated from a single test.

However, despite significant advances in library prep kits, an NGS run can prove labor intensive for even the most skilled technicians. Furthermore, for laboratories that are new to NGS, building the technological skill and proficiency required to achieve consistent and reproducible results takes a considerable amount of time.

Agilent Technologies are working to increase the utility of NGS in the diagnostics field and in personalized medicine. We recently spoke with Ronda Allen, Head of R&D at Agilent, to learn more about two of Agilent's newest products in this space, the Cancer All-in-one (AIO) panels, and the Magnis NGS Library Prep System.  

Molly Campbell (MC): Why are we seeing the increased use of Next Generation Sequencing (NGS) as a diagnostic tool in personalized medicine?

Ronda Allen (RA): The ability to profile many biomarkers in one NGS test has the potential to answer many questions concurrently. This ability is very beneficial in the clinical space as time and tissue biopsies are precious. With continuously decreasing sequencing costs, NGS is becoming more cost-effective than the conventional multimodal sequential molecular testing approach.

MC: What challenges do scientists face when conducting NGS library prep? Please can you tell us how the Magnis NGS Library Prep System has been developed to overcome these challenges?

RA: Even today with advances in NGS library prep kits, the protocol still requires a full-day or multi-shift process by a skilled technician and can be quite labor intensive. For labs that are new to NGS, it can require significant time and investment to establish such expertise, especially in a clinical setting. Even an experienced lab technologist may see quite a bit of variation in the libraries generated using manual protocols due to the complexity of the assay. The Magnis NGS Prep System is an automation system designed to minimize manual steps that could introduce contamination and processing variations. The Magnis requires minimal staff and expertise to run. Agilent provides pre-aliquoted reagents that are barcoded, and the on-board wizard walks users through the set-up in as little as five minutes. This system allows NGS naïve users and labs to start running assays quickly and reproducibly.

MC: Increasingly, we are seeing laboratories move to automated systems for NGS library prep. Why is this, and how can the Magnis NGS Library Prep System help facilitate research for these groups?

RA: As more laboratories adopt NGS for testing, and the volume of tests increases, it becomes critical to have operational efficiency targets. Since the NGS library prep is a time and skill intensive process, well designed automation is the key. The Magnis was designed to run the preparation process in a complete walk-away fashion. Perhaps the most efficient use of Magnis is for a technologist to load the system before the day ends. The preparation completes overnight without intervention. The technologist can then carry on the with the next steps in the morning. Some automation solutions require a technologist to constantly work with the equipment and monitor the progress to ensure reproducibility. Magnis provides the consistency and operational efficiency needed by being a completely walk-away automation solution.

MC: How can the Agilent Magnis NGS Prep system be integrated with other Agilent products to achieve an efficient NGS workflow?

RA: Magnis is already a system of integrated Agilent solutions – from the built-in thermal cycling capability to the single-use packaged SureSelect library preparation and hybrid-capture reagents. In addition, the SureSelect Enzymatic Fragmentation kit is fully compatible as an upstream process to shear the DNA. When a run is completed, the Magnis 8-well library and QC strips can be placed on the TapeStation 4150 for assessing the quality of the libraries. Once sequencing is completed, our Alissa Align & Call and Interpret bioinformatics systems are optimized to analyse SureSelect libraries.

MC: Agilent's Cancer All-In-One (AIO) Panels enable high-sensitivity detection of somatic single nucleotide variants, indels, CNVs and translocations in a single assay. How is this achieved?

RA: This performance is achieved by combining innovative probe design methodology and advanced variant detection algorithms. The AIO panels come with baits designed for the specific variant types that are targeted; the new AIO CNV algorithm tackles the challenge of somatic CNV by enabling clonal CNV detection in samples with low tumor content. Detection of translocations is achieved through the capture and identification of sequences that span the gene fusion point. This approach provides robust translocation calling regardless of the translocation partners.

MC: The AIO panels supports low-input DNA and FFPE samples. Why is this feature beneficial for modern oncology labs?

RA: In current standard practice, clinical samples are preserved as formalin-fixed, paraffin-embedded (FFPE) tissues, which can maintain cell morphology for histological examinations for decades. However, this preservation can cause damage to the DNA, making it challenging for downstream molecular testing. As clinical biopsies are increasingly less invasive, the amount of tissue from surgical biopsies is increasingly small. Thus, it is crucial that an NGS technology for oncology labs performs robustly with small amounts of DNA from FFPE samples.

MC: How does Agilent envision the Magnis NGS Prep System and the Cancer AIO Panels products advancing the field of personalized medicine, specifically in cancer?

RA: The battle against cancer is one that benefits from having more researchers and clinicians join the fight. First, this is enabled by making a powerful technology like NGS available for any lab to run. One of the largest pain-points in NGS is library preparation, and Magnis is the solution to automate that process and deliver consistent results. Second, the field needs powerful assays to ask the right questions. SureSelect has always been at the forefront of library preparation and content selection – the new AIO capabilities provide advanced detection capabilities for both experts and novices alike. We envision that the many experts in oncology – not solely experts in NGS – can now leverage Agilent’s Magnis and AIO technologies to advance better health outcomes.

Ronda Allen, Head of R&D, Agilent was speaking with Molly Campbell, Science Writer, Technology Networks.